Design and Optimization of an IgG Avidity ELISA Based on Toxoplasma gondii Lysate Antigen for Differentiating Acute and Chronic Infections in Pregnant Women

Abstract

Background: Toxoplasmosis is a significant global health concern. Definitive diagnosis of toxoplasmosis is a serious challenge due to false positive serological results. This study focuses on optimization and standardization of an in-house IgG avidity ELISA for Toxoplasma gondii using freshly prepared Toxoplasma lysate antigen (TLA). Methods: Samples were collected during routine pregnancy screening. Following tachyzoite purification, TLA concentration was determined by the Bradford assay at 20 μg/mL. The indirect ELISA assay was then performed with a urea treatment step (6 M). Using 75 serum samples classified as acute or chronic toxoplasmosis, the IgG avidity test was performed by ELISA in two rows of microplates with 6 M urea and PBS, and then the avidity index was calculated. Results: After calculating the cutoff values, the sensitivity and specificity of the in-house Toxoplasma IgG avidity kit were 92% (95% CI: 74–99%) and 100% (95% CI: 86–100%), respectively, with an area under the ROC curve (AUC) of 0.977 (95% CI: 0.951–1.000). Conclusion: The results of the IgG avidity test designed in this study along with standardizing the method and interpreting the results showed the promising performance of this assay to differentiate the acute from the chronic stages of toxoplasmosis